Trigonelline, one of the alkaloids contained in coffee, is important not only as one of the constituents of aroma and flavor in coffee but also as a useful source of nutrition. Its anti-microbial, anti-carcinogenic, and anti-hyperglycemic effects have been investigated in previous studies. However, there have not been any studies examining the anti-degranulation effect of trigonelline. In this study, the anti-degranulation effect of trigonelline was evaluated in in vitro and in vivo models using a rat basophilic leukemia cell line, RBL-2H3 cells, and a passive cutaneous anaphylaxis (PCA) reaction in mice, respectively. In the β-hexosaminidase release assay, trigonelline effectively suppressed antigen-induced degranulation of RBL-2H3 cells in a dose-dependent manner without cytotoxicity. Trigonelline also inhibited FcεRI-mediated intracellular signaling pathways, such as phosphorylation of PLCγ1, PI3 K, and Akt, in antigen-stimulated RBL-2H3 cells and suppressed the PCA response in mice. Moreover, trigonelline also inhibited the microtubule formation in RBL-2H3 cells, indicating that trigonelline could inhibit IgE-sensitized mast cell degranulation by attenuating both the intracellular calcium-dependent and independent pathways. These results revealed that trigonelline possesses the anti-degranulation effect against the development of allergic diseases.